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Isothermal Amplification of Microbial Genomic Samples

DNA based microbial detection and identification assays generally have sample input requirements which are determined by the assay system. In some cases, samples may lack sufficient amounts of microbial genetic material for analysis. If the microbial sample being analyzed is not, or cannot be cultured in the lab, then some means of amplifying the genetic material is required. This is particularly important if multiple assays are required from a single sample. Ideally, the amplification method should amplify all sequences with equal frequency (uniform effect), be generic (applicable to all types of microbial targets), robust (applicable to a wide variety of different sample types) and should have no negative influence on downstream analysis. During development of a new microarray design for microbial fingerprinting, it became apparent that for some samples of interest, sufficient quantities of purified DNA test material were not available, thus an amplification methodology was required. Prior to this study, no commercial off the shelf (COTS) genomic amplification kit was explicitly designed for analysis of microbial samples. Consequently, a diverse set of genomic DNA samples from a number of bacterial agents were amplified using a COTS kit designed for use with human DNA with the presumption that this kit might be useful for microbial samples. The amplified samples were analyzed on a genomic fingerprinting microarray assay platform developed by DRDC. Non-amplified data were co